Anthrax is a well-known infectious disease caused by a Gram-positive bacterium, Bacillus anthracis (B. anthracis). Among the three types of anthrax infection (cutaneous, gastrointestinal, and inhalation), cutaneous anthrax is the most common and is relatively easily treatable with various antibiotics. The other two types of anthrax infections are rare, but usually fatal even with aggressive anti-microbial therapy.
The major virulence factor, anthrax toxin, is composed of three proteins: protective antigen (PA, 83 kilo Dalton, kDa), edema factor (EF, 89 kDa), and lethal factor (LF, 90 kDa). The toxin components act in the binary combinations of PA+EF (edema toxin), and PA+LF (lethal toxin). PA is a cell receptor-binding protein and delivers the other two proteins (EF and LF) into the cytosol of infected cells.
The most effective known method for preventing anthrax is vaccination. The current and only FDA-approved anthrax vaccine in the United States (produced by Emergent BioSolutions Inc. under the trademark BIOTHRAX® (Anthrax Vaccine Adsorbed)) is produced from a sterile cell-free filtrate from an avirulent B. anthracis V770-NP1-R strain. The licensed anthrax vaccine is also called Anthrax Vaccine Adsorbed (or AVA). The vaccine primarily consists of PA, and aluminum hydroxide is used as an adjuvant. The vaccine was developed during the 1950s and 1960s and is licensed by the FDA to Emergent BioSolutions Inc. The vaccine shows less than 0.06% systemic reactions. The ability of the vaccine to elicit an immune response in humans is well-documented. The AVA vaccine is currently licensed for five doses over 18 months followed by annual boosts.
Although the AVA vaccine is effective and safe, new immunogenic compositions for preparing a vaccine that protects a subject against a lethal B. anthracis infection using recombinant technologies are under development. Because protective antigen (PA) is the common factor required for both the actions of LF and EF, it is often used to prepare vaccines for anthrax. Examples of PA vaccines in development include those disclosed in U.S. Pat. Nos. 6,316,006 and 6,387,665 and patent applications US 2010/0183675, US2011/0229507 and WO2010/053610.
Vaccines such as an AVA and PA typically contain at least one adjuvant to enhance a subject's immune response. Aluminum salt adjuvants, frequently referred to as alum, are currently the most widely used adjuvant for use in humans. Alum is usually aluminum hydroxide (also marketed as ALHYDROGEL® (aluminum hydroxide) or aluminum phosphate). AVA and the “next generation” Anthrax vaccines (such as recombinant PA) are formulated with aluminum hydroxide which binds the antigen.
Currently, vaccines containing alum require a cold chain. Cold chains have been established globally to keep vaccines at 2-8° C. during storage and distribution. Maintaining cold chains is expensive and difficult. In the event of a cold chain failure, vaccines can be exposed to higher or lower temperatures than intended. It is generally recommended that vaccines that contain alum be discarded if they undergo freeze/thaw processing during shipping and storage. Failure of a cold chain can occur in both industrialized and developing nations, and there are many reasons for cold chain failure, for instance, equipment failure, lack of resources or poor compliance. In many developing countries such as Indonesia, freezing temperatures were recorded in 75% of baseline shipments and freezing of freeze-sensitive is widespread. See Hepatitis B vaccine freezing in the Indonesian cold chain: evidence and solutions. Bulletin of the World Health Organization 2004; 82:99-105.
A vaccine that is dependent on a cold chain may also take longer to distribute to those in need in a timely manner. In the event of a bioterrorist event or other public health emergency, the ability to rapidly deliver vaccines and other medical countermeasures is critical. Eliminating dependence on the cold chain for distribution would lead to more prompt and efficient delivery of medical countermeasures in a variety of climates.
In order to avoid or minimalize cold chain requirements, many licensed vaccines are formulated as a dry powder composition that can be reconstituted immediately prior to administration. To date, all dry powder vaccines licensed for use in the US are produced through a lyophilization process. Lyophilization, also referred to as freeze drying, is a process that improves the long term stability of a vaccine. The process involves freezing the liquid vaccine formulation and subliming the frozen formulation under vacuum. Other technologies such as spray drying and foam drying have been developed with the aim of producing a stable, dry powder vaccine. These newer technologies produce dry powder vaccine material without the need for freezing and can be used with an alum containing vaccine. See, for instance, Chen et al., 2010, Vaccine 28:5093-5099. However, these newer technologies are still in their infancy and have yet to be used in the production of a licensed vaccine in the United States.
Freezing of vaccine compositions containing alum (either as part of the lyophilization process or to produce a frozen vaccine) generally induces aggregation of the aluminum particles and causes degradation of the antigen adsorbed onto the alum adjuvant resulting in potency loss. In addition, freezing causes reduction of the height of the settled aluminum gel (commonly referred to as gel collapse). See, for instance, “The effect of freezing on the appearance, potency and toxicity of adsorbed and unadsorbed DPT vaccines,” 1980, WHO Weekly Epidemiological Record 55:385-92; “Temperature Sensitivity of Vaccines,” August 2006, WHO publication WHO/IVB/06.10; Diminsky et al., 1999, Vaccine 18(1-2):3-17; Maa et al., 2003, J Pharm Sci 92(2):319-332.
Accordingly, there is a need to produce a vaccine that contains alum that can withstand freezing. Such a vaccine may be subjected to freezing as part of the manufacturing process (e.g., a lyophilized or frozen vaccine), shipping process or during storage. The present invention discloses novel formulations for the production of temperature stable vaccines containing alum.